Membrane for an in-vitro resorption model of the gastrointestinal tract

ABSTRACT

A porous membrane adapted for use as the filter element in an in-vitro resorption model of the gastrointestinal tract, said membrane consisting essentially of solid porous carrier material, especially porous cellulose nitrate, impregnated with a liquid phase consisting of a mixture of a higher fatty acid and a neutral lipoid component, particularly a mixture of lauryl alcohol and caprylic acid.

States Patent Herbert Stricker Inventor lngelheim/Rhine, Germany App]. No. 861,777 Filed Sept. 29, 1969 Patented Oct. 12, 1971 Assignee C. H. Boehringer Sohn Ingelheim/Rhine, Germany MEMBRANE FOR AN IN-VlTRO RESORPTION MODEL OF THE GASTROINTESTINAL TRACT 5 Claims, 2 Drawing Figs.

US. Cl 210/490, 21Q/506 int. Cl B01! 39/14, BOld 31/00 Field of Search 210/22, 23,

[56] References Cited FOREIGN PATENTS 17,503 8/1915 Great Britain 210/23 OTHER REFERENCES Ferry, Ultrafilter Membranes and Ultrafiltration, from Chemical Reviews, vol. l8, 1935, pp. 373- 455, pp. 410- 412 relied on.

Primary ExaminerFrank A. Spear, Jr. Atrorney-Hammond & Littell STRACT: A porous membrane adapted for use as the filter element in an in-vitro resorption model of the gastrointestinal tract, said membrane consisting essentially of solid porous carrier material, especially porous cellulose nitrate, impregnated with a liquid phase consisting of a mixture of a higher fatty acid and a neutral lipoid component, particularly a mixture of lauryl alcohol and caprylic acid.

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BNVENTOR HERBERT STRICKER a ,(rdz. (-13 ATTORNEYS 3,bl2,2h3

MElt/ihlliitifllii li Olli AN llhl-VllTTTO EESOIRWHON MODEL OlF TliillE GASTMOTNTEETIINAL 'lflitliiCT BACKGROUND OF THE llNi/ENTTON The effectiveness of a drug in the system of the human body is, inter alia, a function of its concentration at the place of effective action and thus also a function of its ability to reach that place. Most of the perorally administered drugs must first overcome the barriers of the gastrointestinal tract, i.e., they must be resorbed, before they can be transported by the blood to the place where they exert their therapeutic action.

Various in-vivo methods for following the passage of chemotherapeutics from the gastrointestinal tract into the blood circulation system are known.

For instance, the decrease in the concentration of the chemotherapeutic substance with passage of time in various parts of the gastrointestinal tract after peroral administration can be determined by analytical methods in the tied-off stomach, colon, or small intestine of rats.

In humans, the change in concentration of various dissolved organic acids and bases in the stomach over a period of time can also be followed by means of an esophageal sound lowered into the stomach.

Likewise, in humans as well as animals, the blood level values at various times after peroral administration of drugs can be determined, from which rate of resorption can then be calculated.

Such in-vivo methods, however, require a substantial expenditure of work and money, and the results obtained therefrom are often subject to wide variations with respect to accuracy and reproduceability.

ln-vitro methods employing artificial membranes, on the other hand, represent a valuable supplementation of the invivo determinations. Thus, by means of greater series tests and more accurate concentration determinations, the kinetics of diffusion can be very accurately ascertained, so that the effects of the interchange action between the active ingredients among themselves or with the pharmaceutical excipients can already be evaluated and taken into consideration in determining the most favorable makeup of the pharmaceutical dosage unit compositions. Moreover, by employing in-vitro methods, the number of in-vivo tests can be significantly reduced.

THE PRlOR ART lFilter membranes for various purposes have already been described in the literature. For instance, porous membranes made of cellulose esters and impregnated with olive oil or linoleic acid have been described by G. Levy and E. J. Mlroszezak in J. Pharm. Sci. 57(2), 235-239 (1968).

However, it was found that, while these olive oil impregnated filter membranes are pl-l impermeable," they also do not permit the passage of substances of an acid character if they are present in weakly acid to neutral solution. Therefore, the olive oilimpregnated membranes are not well suited for in-vitro resorption tests, especially if the apparatus is to simulate resorption in the small intestine.

On the other hand, linoleic acid-impregnated membranes proved to be insufficiently permeable with respect to hydrogen or hydroxyl ions, so that they could not be used for diffusion measurements between solutions having a difierent pH value.

OBJECTS OF THE llNi/ENTTON The present invention has as its principal object to provide a membrane for use in an in-vitro resorption test apparatus, which is capable of maintaining the pH gradient between two aqueous solutions substantially constant over a period of time of about 50 hours and exhibits a specific permeability for acid, basic, and amphoteric drug molecules.

Another object of the instant invention is to provide a porous membrane for use in an in-vitro resorption test apparatus, which duplicates as closely as possible the drug diffusion rates under physiological conditions known from in-vivo tests.

Other objects and advantages of the invention will become apparent as the description thereof proceeds.

THE llNVENTlON The above objects are achieved in accordance with the present invention by impregnating a. solid, porous, carrier material with a liquid lipoid phase.

The porous carrier material may be a commercial membrane filter material made of a cellulose ester. The diameter of the pores must be sufficiently small so that the pores, due to their capillary activity, remain closed by means of the liquid impregnation component; otherwise, the liquid impregnation phase could be dissolved out and the membrane would become useless. Therefore, the diameter of the pores should be from 0.05 to 1.0 mg, preferably 0.1 mg. Examples of suitable membrane filter material are the commercially available products lviillipore VC and Sartorius SM 1 1309.

The liquid lipoid phase must resemble with respect to its physical properties, such as its relative solution capacity for acid, basic, and amphoteric drugs, and its distribution as closely as possible to its natural counterpart, i.e., the stomach and intestinal walls, so that the relative resorption rates of various drugs can be accurately simulated in the in-vitro apparatus.

l have found that these requirements are met by a mixture consisting of a higher fatty acid, preferably caprylic acid, and a neutral lipoid component, such as a higher fatty alcohol of 12 or more carbon atoms, preferably lauryl alcohol. The mixture of these two components must be liquid at the test temperature of 37 C.

A membrane according to the present invention is prepared by immersing the porous filter material into a liquid mixture consisting of a higher fatty acid and a neutral lipoid component, removing the impregnated porous material from the liquid mixture and allowing excess liquid mixture to drip off, and blotting the impregnated porous membrane between filter paper until its weight is constant. A preferred embodiment of the membrane is obtained by using as the liquid-impregnating component a mixture of parts by volume caprylic acid and 20 parts by volume lauryl alcohol.

in contrast to known membranes of this type, membranes in accordance with the present invention have the advantage that they are capable of maintaining the pll-ll gradient between two aqueous solutions substantially constant (:c about 0.3 pH units) over a period of about 50 hours and exhibit a specific permeability for acid, basic, and amphoteric drug molecules. The diffusion rates under physiologicai conditions correspond substantially to the values known from in-vivo tests.

The combination of a higher fatty acid and a neutral lipoid component with a cellulose ester filter membrane as a carrier material proved to be a system which meets the requirements surprisingly well. The effect of the fatty acid component is that, corresponding to its pit, value and the pit-ll of the adjacent solution, it is partly present as an anion and acts as a proton acceptor or donor with respect to the dissociated organic acids or bases. The ,drugs are thereby transformed at the membrane surface into the undissociated form and are thus capable of migrating through the liquid phase of the membrane, whereas the hydrogen and hydroxyl ions can pass through only very slowly because of the solubility conditions.

The annexed drawings show schematically the type of invitro diffusion apparatus wherein a membrane according to the present invention can be used.

FIG. 11 is a schematic representation of an apparatus which consists essentially of two chambers i and la (volume of each, ml.) which are separated by a membrane 2 according to the present invention (effective surface 12.5 cm") Each of the chambers is provided with a thermostatically controlled exterior heating jacket 3 to maintain the contents at 37 0%.5" C. Two pipettes d and 5 extend into each of the chambers T and lla, and a dropping funnel (not shown) is connected to the exterior end of .each pipette. The flow of liquid through each pair of pipettes is controlled by a double stopcock 6. Pipettes terminate substantially at the point where they enter chambers 1 and 1a, respectively, whereas pipettes 4 extend into the respective chambers to a point close to the surface of membrane 2. One of the two chambers 1 and 1a is filled with a physiological solution having a pH of 7.5, which corresponds to that of blood plasma; the other chamber is filled with a physiological solution whose pH is adjusted to between 1.2 and 7.0, depending upon the section of the gastrointestinal tract which it is intended to duplicate. The dropping funnel connected to each of the short pipettes 5 is filled with buffer solution to maintain the pH in each chamber at the desired level. A metal disc 7 having a central aperture is slidably mounted on each pipette 4. The entire apparatus is mounted so that it oscillates 36 times per minute through an angle of 170 about a transverse axis 8 which coincides with the center of the plane formed by membrane 2, the oscillating motion being provided by a motor (not shown). By virtue of this oscillating motion the metal discs 7 slide along pipettes 4 and thereby stir the solution in each chamber. In order to withdraw a sample of the solution in chamber 1 or la, the respective double stopcock 6 is opened, whereby the descending buffer solution in pipette 5 forces some solution from the chamber upward through pipette 4 into the dropping funnel connected thereto, wherefrom it can be withdrawn for analysis. At the same time the solution in the chamber is replenished with fresh buffer solution. The function of membrane 2 is to maintain the pH gradient between the solutions in chambers l and 1a, and also to permit differential passage of various drugs, i.e., organic acids and bases of different types. The drug whose diffusion rate is to be tested is introduced into the solution which duplicates the particular section of the intestinal tract.

FIG. 2 is a schematic representation of another diffusion apparatus which comprises a diffusion vessel 9 consisting of two chambers 10 and 10a separated by a membrane 11 according to the present invention. The test solutions are cycled by means of pump 12 from storage vessels l3 and 130 into chambers 10 and 10a, respectively, and then back into the storage vessels. The cycling rate depends on the dimensions of the apparatus; for example, if the membrane surface is about 12 cm. and the volume of chambers 10 and 10a is about 2 to 5 ml. each, the cycling rate is advantageously about 2 to ID ml./min. In addition, the solutions in storage vessels 13 and 13a are continuously stirred by means of a magnetic stirrer 14 in order to avoid concentration gradients. Thermostatically controlled heating jackets l5 maintain a constant temperature of 37 C. Stopcocks l6 permit withdrawal of samples of the solutions for analysis.

The following examples further illustrate the present invention and will enable others skilled in the art to understand it more completely.

EXAMPLE 1 Using a diffusion test apparatus such as the one shown in FIG. 1 of the drawings, the resorption rate of various acid and basic drugs from the stomach was tested.

The membrane was a porous sheet of cellulose nitrate, pore size 100i8 my. (Millipore VC or Sartorius SM 11309), impregnated with a liquid mixture consisting of 50 parts by volume of caprylic acid and 50 parts by volume of lauryl alcohol.

One difiusion chamber was filled with a solution of the drug compound in 100 ml. of a buffer solution having a pH of 1.2, i.e., the pH prevailing in the stomach, prepared from 24 g. of l N HCL, 0.5 g. of glycocoll, 0.35 g. of NaCl and enough H O to make 1,000 ml.

The other diffusion chamber was filled with 100 ml. of a buffer solution having a pH of 7.5, prepared from 20.5 g. of Na HPO,, 2.7 g. of KH PO and enough H O to make 1,000 ml.

The apparatus was oscillated as described above in connection with FIG. 1 of the drawings, and samples of the solution in each chamber were withdrawn periodically, while at the same 5 time replenishing the buffer solution from the dropping funnels.

The following table shows the results obtained in comparison to known in-vivo resorption rates, all values being based on the resorption rate of aspirin=l .00.

TABLE I 15 Initial resorption rate, based on rate of ace! lsalic lic acid=l.00

Compound in vivo' in vitro Acetylsalicylic acid 1.00 1.00

Salicylic acid 2.3 2.3 Bcnzoic acid 2.0 2.2 Phenol Red 0.05 0.1 Aminopyrine 0.05 0.0 Quinine 0.00 0.0 Ephedrine 0.00 0.0

' L. S. Schanker et 21.,1. Pharm. exp. Thcrap. I20, 528 53 EXAMPLE 2 The test procedure and apparatus were the same as in example 1, except that the membrane was impregnated with a liquid mixture consisting of 20 parts by volume of lauryl alcohol and 80 parts by volume of caprylic acid, and the buffer solution having the drug compound dissolved therein had a PH of 6.0 to simulate conditions in the small intestine; the buffer solution was prepared from 1.5 g. of Na HPO 80 g. of KH PO and enough water to make 1,000 ml. The following results were obtained:

' L. S. Schanker et al.,J. Pharmakol. exp. Therap. I23 8] (I958 The results of these examples clearly show that, with the aid of an in-vitro apparatus comprising a membrane according to the present invention it is possible to duplicate very closely the in-vivo resorption conditions of the gastrointestinal tract.

Iclaim:

l. A membrane for an in-vitro resorption model of the gastrointestinal tract, said membrane consisting of a porous cellulose ester filter membrane material having a pore size of 0.05 to 1.0 mg. impregnated with a mixture consisting of a higher fatty acid and a higher fatty alcohol, said mixture being liquid at 37 C.

2. A membrane according to claim 1, wherein said liquid mixture consists of caprylic acid and lauryl alcohol.

3. A membrane according to claim 1, wherein said mixture consists of 20-50 percent by volume of lauryl alcohol and 5. A membrane according to claim 1, wherein said filter membrane material is porous cellulose nitrate having a pore size of about 100 mp, and said liquid mixture consists of 20 percent by volume of lauryl alcohol and percent by volume of caprylic acid.

"H050 UNITED STATJ'JS PATENT OFPICE 1 1 1 'fif ERTIFICA'HL O1 CORLILL I lON Patent No. 3,6 3 Dated October 12, 1971 Inven t o r M HERBERT STRICKER It is certified that error appears in the above-:Ldentified patent and that said Letters Patent are hereby corrected as shown below:

ln the first page, insert [321 Priority p 7 6 signed and 365116.11 this 28th d 'a of Maro.h 'l972.

(SEAL) Attest:

EDWARD M.FLETCHER,JR. p ROBERT GOTTSCHALK {it-testing Off1cer Commissioner of Patents 

2. A membrane according to claim 1, wherein said liquid mixture consists of caprylic acid and lauryl alcohol.
 3. A membrane according to claim 1, wherein said mixture consists of 20-50 percent by volume of lauryl alcohol and 50-80 percent by volume of caprylic acid.
 4. A membrane according to claim 1, wherein said filter membrane material is porous cellulose nitrate having a pore size of about 100 m Mu , and said liquid mixture consists of equal parts by volume of caprylic acid and lauryl alcohol.
 5. A membrane according to claim 1, wherein said filter membrane material is porous cellulose nitrate having a pore size of about 100 m Mu , and said liquid mixture consists of 20 percent by volume of lauryl alcohol and 80 percent by volume of caprylic acid. 